Project Summary This K99/R00 career development award proposal describes a two-year mentored and three-year independent research program essential for the development of the principal investigator as an independent investigator. The principal investigator received his Ph.D. in Neuroscience at the Johns Hopkins University for his work on the differential regulation of pain and itch sensations by a small subset of spinal cord interneurons, and the molecular mechanisms by which proteases induce itch sensation in the periphery. This research project was conducted under the supervision of Dr. Xinzhong Dong. He then joined Dr. Ulrich Mueller's laboratory at Johns Hopkins to receive training in auditory neuroscience. His specific project focused on the analysis of the mechanisms that regulate the differentiation of auditory sensory afferent neurons. The study revealed neuronal heterogeneity in auditory afferents and demonstrated that activity dependent mechanisms are critical for neuronal diversification. The proposed training for this K99/R00 application will be supported by an advisory team consisting of Dr. Mueller (mentor), Dr. Sascha du Lac (co-mentor) and Dr. Loyal Goff (consultant). The advisory team will assist with specific techniques, help to identify job opportunities, aid in preparation for job interviews and help to build his laboratory. In addition, proposed courses, scientific meetings and seminars will complement the training program to accomplish his goal to become a successful independent investigator. The long-term research goal of the proposed work is to define the molecular mechanisms by which vestibular circuits differentially encode sensory information to shape our sense of balance. Specifically, this proposal will focus on the vestibular ganglion neurons (VGNs), which transmit vestibular information to the CNS. The research plan proposes to test the central hypothesis that VGNs are a group of functionally diverse neurons that can be categorized into subtypes which transmit distinct aspects of vestibular information to the CNS. This hypothesis is built upon preliminary data showing that 1) VGNs consist of molecularly distinct subtypes defined by single- cell RNA sequencing (scRNAseq) and verified at RNA and protein levels; and 2) activity-dependent mechanisms contribute to the establishment of VGN circuits. During the mentored phase, the proposed studies will define the molecular heterogeneity of VGNs; characterize VGN subtypes; and map their central and peripheral projections. In the independent phase, the investigator will built on the data collected in the mentored phase to investigate functional and developmental diversification of VGN subtypes. The central hypothesis will be tested using an experimental strategy that combines scRNAseq, genetic based circuit mapping, functional circuit perturbation and quantitative behavioral analyses. The results will lay a solid foundation to support the principal investigator's research goals that hold the promise to offer better understanding of 1) the functional diversity of the molecularly distinct cell types in the vestibular system; 2) the genetic deterministic programs and activity-dependent mechanisms in circuit formation and function; and 3) circuit maladaptation in aging and disease.